Iron dextran complex and process for making same

ABSTRACT

A method of making iron dextran complexes wherein ferric hydroxide is reacted with dextran or dextran glucosides, precipitating iron dextran into at least two fractions having different iron contents and subjecting one of the fractions of the iron dextran formed in the first complexing step to a second complexing step and precipitating the iron dextran into at least two fractions having different iron contents, separating and recovering the fraction having the higher iron content and recycling the fraction having the lower iron content.

Unite States Patent de Musquiz Reumert et al.

[1 1 3,666,749 51 May 30, 1972 [54] IRON DEXTRAN COWLEX AND PROCESS FORMAKING SAIVIE [72] Inventors: Jorgen de Musquiz Reumert, Gentofte; OleGylling-Pedersen; Jose Bou Casals, both of Roskilde, all of DenmarkAktieselskabet Rosco, Taastrupgardsveg, Taastrup, Denmark 22 Filed:Sept. 15,1969

21 Appl.No.: 858,105

{ 73] Assignee:

Related US. Application Data [63] Continuation-in-part of Ser. No.502,485, Oct. 22,

I965, abandoned.

[30] Foreign Application Priority Data Dec. 29, 1964 Denmark ..6403/64[52] US. Cl. .260/209 D, 424/180 [51] ..C07c 47/18 [58] Field of Search..260/209 D [56] References Cited UNITED STATES PATENTS R24,642 4/1959London at al ..260/209 D 2,885,393 5/1959 Herb ..260/209 D 3,093,5456/1963 Westfall et a1. ..260/209 D Primary Examiner-Lewis GottsAssistant Examiner-Johnnie R. Brown Attorney-Synnestvedt & Lechner [5 7]ABSTRACT 9 Claims, No Drawings IRON DEXT'RAN COMPLEX AND PROCESS FORMAKING SAME This application is a continuation-in-part of our US. Pat.application, Ser. No. 502,485 filed Oct. 22, 1965 and now abandoned.

The present invention relates to the manufacture of compositionscomprising a substantially nonionic complex of ferric hydroxide with adextran.

Preparations for injection of aqueous solutions of complexes of ferrichydroxide with a dextran have been extensively used in prophylactic andtherapeutic treatment of iron-deficiency anaemia. Several processes areknown for producing such complexes, all being based on the reaction offerric hydroxide with a dextran.

Dextran is a polysaccharide with the empirical formula (C l-i and it isbuilt up of glucose units being linked together mainly through alpha-l.6-glucosidic bindings and to a considerably less extent throughalpha-lA-gluoosidic bindings.

In the known processes for producing the complexes which may be callediron dextran complexes, partially depolymerized dextran or raw dextranwith a relatively small average molecular weight and a ferric compoundare used as starting material. In addition thereto use is made of abase.

Thus, a process is known in which partially depolymerized dextran isadded to a solution or a suspension of a ferric compound whereafter abase is added for the formation of ferric hydroxide, or the dextran isdissolved in alkali whereafter the solution or the suspension of theferric compound is added and the ferric compound is converted intoferric hydroxide. It is preferred to add a solution of the base to anaqueous solution of the dextran and thereafter to add an aqueoussolution or suspension of the ferric compound. In the said process thereaction or combination of ferric hydroxide and dextran is carried outat a temperature between room temperature and the boiling point of thesolution, and the operation takes place at a pl-l-value within the basicrange. A dextran having an intrinsic viscosity within the range of 0.025to 0.5, particularly 0.03 to 0.06, is used. After purification of theprepared product iron preparations being isotonic with the blood fluidscan be produced. A process of this kind is described in the British Pat.specification, No. 748,024 and the US Pat. specification Re 24,642.

According to another known process a dextran decomposition producthaving a molecular weight within the range of about 30,000 to 80,000 isused as starting material. Also in this case there is in addition to thetwo reactants, dextran and ferric compound made use of a base to formferric hydroxide, and the reactants may be added to each other in anysequence. If the dextran is added to a ferric salt solution prior to theaddition of the base, it is not necessary to control the addition of thebase. if, on the contrary, the base is added to the ferric salt solutionbefore the dextran is added, the pH-value should be adjusted to at leastabout 2.0, preferably to about 2.3. After the complexing reaction, whichcan be accelerated by heating, the complex may be isolated and purifiedby repeated precipitations and re-dissolving in water. It is preferredto heat the complex to partially depolyrnerize the dextran, and thesolution is made alkaline and is heated again, which causes allunreacted iron to be taken up by the dextran. Thereafier the solution isneutralized and the complex of ferric hydroxide with dextran isisolated. The isolated complex, which may show an iron content withinthe range of 18 percent to about 27 percent by weight, is then dissolvedin water to form a solution which can be brought to any desiredconcentration and which is made isotonic with the blood fluids. it is,however, not possible to have the iron content exceed 7 percent. Aprocess of this kind is described in US. Pat. specification, No.2,885,393.

Further, it is known to prepare an injectable iron dextran complex byheating a dextran glucoside having an average molecular weight of 500 to80,000 and a fem'c salt to a temperature between 100 and 120 C in thepresence of an alkali hydroxide. The resultant reaction product betweenferric hydroxide and the dextran glucoside is purified and dried, ifdesired, whereafter a solution being isotonic with the blood fluids canbe prepared.

There is also known a process according to which a complex of ferrichydroxide with dextran is produced from raw dextran having an intrinsicviscosity at 25 C between 0.25 and 0.75, by heating a solution or asuspension of the raw dextran with a solution of an acid ferric saltuntil the intrinsic viscosity of the dextran is not more than 0. l at 25C, whereafter the said mixture is treated with aqueous alkali hydroxide.Then the resultantcomplex is isolated or purified in the usual manner.The purified complex contains 22 to 27 percent of iron by weight. Fromthe complex it is possible to prepare sterile solutions being isotonicwith the blood. This process is described in the German Pat.specification, No. 1,172,250.

Finally, it is known to produce a complex of ferric hydroxide withdextran by adding to an acidic solution of water-soluble dextran and awater-soluble iron compound an aqueous solution of an alkali metalcarbonate or bicarbonate, subsequently adding an alkali metal hydroxideto form a suspension, heating said suspension to fonn a solution andneutralizing the solution to obtain a complex of ferric hydroxide withdextran, said complex containing about 33% Fe by weight and showing aratio of 2 moles of iron to each anhydroglucose unit. This process isdescribed in the SwissPat. specification, No. 370,194.

The present invention relates to a process by which it is possible notonly to arrive at iron dextran complexes with a high ferric content andeven a higher one than hitherto known, but also to obtain an improvedutilization of the dextran compound used as starting material. Animproved utilization of the employed amount of dextran compound is ofeconomical importance and results in final products with a morefavorable ratio of iron to dextran compound.

In the known processes a freshly prepared solution of ferric hydroxideor a solution of ferric hydroxide formed in situ is reacted with adexu'an solution to form the complex of ferric hydroxide with dextran,and the said complex is isolated by precipitation with a water-misciblesolvent, such as methyl alcohol, ethyl alcohol or acetone, and sometimespurified by redissolution in water and reprecipitation.

The principal feature of the process of the present invention comprisesfractional precipitation of the formed complex of ferric hydroxide witha dextran compound to form at least two fractions showing different ironcontents and utilization of one of these fractions as starting materialinstead of the dextran compound for reaction with ferric hydroxide toform a complex of ferric hydroxide with the fraction used instead of thedexu'an compound. In other words, the process of the invention comprisesa recycling step, and this recycling step makes an improved utilizationof the dextran compound possible whereas it will also become possible toproduce iron dextran complexes showing a higher iron content than thosehitherto known.

By the process of the invention it is possible to arrive at iron dextrancomplexes having an iron content up to about 42 percent of the dryweight, and from the said complexes it is possible to prepare injectablesolutions having an iron content of 12 percent, and such solutions showa low viscosity and LB values between 750 mg and 1,300 mg of iron perkilogram of the weight of the body when injected intravenously in mice.This has been confirmed by the pharmacological testings on sucking pigsduring which no cases of death due to the said ferric dextranpreparation have been encountered.

When in the first step of our process the complexing reaction takesplace between ferric hydroxide in solution and the dextran compound insolution, the dextran compound should show an intrinsic viscosity withinthe range of about 0.0025 and about 0.25 at 25 C. When however, the saidfirst complexing step is carried out under such conditions that also amaterial depolymerization of the dextran compound is obtained, it ispossible to use as starting material dextran compounds having anintrinsic viscosity up to 0.75 at 25 C. The

intrinsic viscosity of the dextran compounds being used as startingmaterial should not be below 0.025 at 25 C since lower values do notyield satisfactory products.

The dextran compounds used in the first step of our process is dextranor dextran glucosides which have the desired intrinsic viscosity. Bothsuch dextran and such dextran glucosides are known in the art. Withrespect to dextran glucosides particular reference is made to US Pat.specification, No. 2,929,764, and among the dextran glucosides describedtherein it is preferred to use dextran glycerol glucoside.

The ferric hydroxide used in our process may be formed from anywater-soluble ferric salt which together with a base in the presence ofdextran forms ferric hydroxide, or use may be made of mixtures of suchferric salts. It does not greatly matter which anion forms part of thesaid salts since the anion does not partake in the reaction. Examples ofsuch salts are ferric chloride, ferric nitrate, ferric sulfate, ferricacetate, ferric trichloro acetate, ferric citrate and double salts, suchas ferric ammonium sulfate and ferric hydroxy salts. Freshly preparedferric hydroxide may also be used.

The basic-reacting compound to create the formation of the ferrichydroxide may be an alkali metal hydroxide, alkali metal carbonate orbicarbeonate, ammonium hydroxide, ammonium carbonate or bicarbonate, ortetramethyl ammonium hydroxide. The amount of base to be added dependson the complexing conditions used, and the preferred base depends on theferric salt used. it is known in the art how to form the ferrichydroxide for complexing with dextran.

The ratio between the dextran or ferric dextran compound used in thecomplexing reaction and the ferric compound may be of importance forachieving the desired product. When e.g., dextran glycerol glucoside isused as dextran starting compound, the said ratio should e.g., bebetween about 1.1 and about 4 moles of anhydroglucose units (C H O inthe dextran compound per mole of iron. it is preferred not to use morethan about 2 moles of glucose units in the dextran compound per mole ofiron.

Though it is possible to carry out the complexing reactions at roomtemperature, increased temperatures up to the vicinity of l00 C areusually used for obtaining a reasonable reaction velocity.

The complexes resulting from the complexing reactions consist likedextran of molecules having difierent sizes and the iron content of thesaid molecules expressed in percentage is varying.

The complexes fonned by the first complexing step are as mentioned abovesubjected to a fractionate precipitation. Thereby first the fraction orfractions having the greatest iron content are precipitated. The saidfractions are usually those having the highest average molecular weight.

in the precipitation step which is known in the art use may be made of awater-miscible organic solvent, such as acetone or lower molecularaliphatic alcohols, e.g., methanol, ethanol or propanols. In view of thefact that the fractionated precipitation will be the more complicatedand consequently more expensive to carry out the more numerous are thefractions into which the product from the first complexing step isprecipitated, it is according to the invention appropriate that theproduct from the first complexing step is precipitated into twofractions having difierent ferric contents, the fraction'having thelower ferric content being used in the second complexing step.

Thefractionated precipitation may be controlled by varying the ratiobetween solvent and precipitant. Consequently, it is possible accordingto ones desire to carry out the precipitation so that the last isolatedfraction will be big and have a relatively small iron content, and thefirst isolated fraction will be small and have a relatively high ironcontent.

The ferric content of the iron dextran complex used as dextran startingmaterial in the second complexing step may vary within large limits.Thus, it is on one hand possible to use a complex with a ferric contentbeing as low as e.g., 0.i per Cent by weighnand on the other hand alsopossible to use an iron dextran complex having an iron content as highas e.g., 22 percent by weight.

According to the invention it is appropriate to use as iron dextrancomplex in the second complexing step a fraction having an iron contentof 0.3 to 3 percent by weight.

Hereby is obtained that the fraction to go on in the process will belarge enough to secure that a high yield of the complex of ferrichydroxide with the dextran compound with a high iron content will beobtained while obtaining at the same time the economic advantage thatthe fraction or fractions first precipitated in the fractionatedprecipitation, being that or those having the greatest iron content, canalso be used for preparing iron dextran preparations though with lessiron content, such as solutions containing about 5 to 7.5 percent ofiron. The average molecular weight of the said fractions should not beso high that iron preparations produced therefrom will be too viscous,which could make them inapplicable in practice.

Further, it is appropriate according to the invention that the productfrom the first complexing step is purified prior to the fractionatedprecipitation. Thereby it is better ensured that the final product willhave the desired low toxicity and besides that the fraction or fractionshaving the highest iron content will be purified.

By the said purification the electrolytic constituents will be removed,first of all ferric ions. This also applies to the purification afterthe second complexing step. Both of these purifications may be carriedout e.g., by dialysis, ion exchange or one or more precipitationsfollowed by re-dissolving.

According to the invention the second complexing step is carried out inan acid medium, since it has been found that this results in theformation of iron dextran complexes having a high iron content.

When carrying out the second complexing step in an acid medium it isaccording to the invention preferred to form the ferric hydroxide froman inorganic ferric salt or an organic ferric salt being strongly acidin solution, e.g., ferric trichloro acetate.

Since, as mentioned above, a complexing reaction will always result in afinal product having molecules of different sizes and molecules havingvarying iron content so that those constituents of the product having ahigh molecular weight will usually have a high iron content, and sincethe said constituents of the product will further be first precipitatedby a fractionated precipitation, it is according to the inventionappropriate that the product from the second complexing step, in which afraction from the first complexing step is used as one of the reactantsinstead of the dextran compound itself, is precipitated into twofractions with different iron contents, the fraction showing the loweriron content being recycled.

By the above measures a final product is obtained which has an elevatedaverage iron content and the final yield is not very much reducedbecause of recycling the fraction having the lower iron content.

The said precipitation should be carried out in such a manner that thetwo fractions will have almost the same size.

If the fractionated precipitation after the second complexing step iscarried out in such a manner that the fraction having the higher ironcontent will be considerably greater than the fraction having the loweriron content this will involve a reduction of the iron content inpercentage in the first mentioned fraction, and solutions thereof willwith the same iron concentration have an undesired high viscosity.

Further, it is appropriate according to the invention to use as startingmaterial in the first complexing step a dextran compound which ispurified and, if desired, isolated and dried. By the said measure it isbetter ensured that the final product will have the desired lowtoxicity.

It is according to the invention appropriate to carry out both of thecomplexing steps at a pI-l-value within the range of 1.1 to 2.3.Preferably a pl-l-value of 1.4 to 1.6 is used. Thereby higher yields ofthe iron dextran complex having a high iron content are obtained.

In the said embodiment a ferric salt is dissolved in water and to thesaid solution a solution of a base is added slowly while stirring. About1.35 to 1.7 moles of base equivalent per mole of iron are used, wherebypH-values within the desired range of 1.1 to 2.3 are arrived at. Theratio between base and ferric compound is fixed in order to secure thatthe ferric hydroxide formed remains in colloidal solution, and thepl-l-value will not be too low or too high to enable the complexing totake place. Besides on the mole ratio between base and ferric compoundthe pH-value is also dependent on the concentration of the solutions.

Thereafter a solution of the dextran compound is added to the colloidalsolution of the partially neutralized ferric salt at an elevatedtemperature, e.g., 40 to 90 C, and the complexing step is effected atthe elevated temperature for a period of time ranging from 5 minutes to2 hours.

Then the complex formed is purified by means of one of the abovementioned purification methods so that the pH-value of the solution willbe between 2.0 and 2.3, which means that the main part of the ferricions has been removed.

Then autoclaving at about 110 to 120 C for to 60 minutes is carried out,which causes the dextran to depolymerize somewhat. The said autoclavingis important for obtaining a satisfactory final product.

After autoclaving and cooling a base, preferably a sodium hydroxidesolution, is added until the pH-value is at least 10 whereafterautoclaving at 120 C for 10 to 60 minutes is again carried out. Thiscauses the last remainders of unreacted ferric ions to be complexed.After cooling the pI-l-value is adjusted to about 6.0 with hydrochloricacid, whereafter the product is precipitated into two fractions showingdifferent iron contents. Both fractions are dried, e.g., in a vacuumunit at about 60 to 70 C. The fraction having the lower iron content isbrought back to the process while the other fraction can be used forpreparing injectable iron dextran solutions.

When carrying out the process according to the said embodiment it isaccording to the invention appropriate to use as dextran compound acompound having an intrinsic viscosity of 0.08 to 0.12. l-lerebycomplexes are obtained which in solutions have a viscosity suitable forinjection.

The complex from the second complexing step can, after it has beenisolated, be brought directly into solution in a suitable,pharmacologically acceptable solvent.

Water is used as solvent. Prior to being used as a therapeuticpreparation the said solution should be made isotonic with the bloodfluids. This may be done in a manner known per se. The pl-l-value of thefinished solution should be within the range of 5.5 to 8.5, andpreferably solutions having a pH-value of 6.7 to 7.5 are produced.

The product from the second complexing step may also be isolated byprecipitation and subsequent drying. The resultant solid iron dextrancomplexes may immediately be dissolved in distilled water for use in theproduction of therapeutically applicable, injectable solutions of theiron dextran complex.

When such a solution has been prepared it may be sterilized prior tobeing poured into the final containers, or it may e.g., be poured intoampoules and then sterilized. When sterilization is carried out beforethe solution is poured into the final containers it is appropriate witha view to securing the sterility of the solution to add a preservingagent.

The said iron dextran complexes cannot only be used in the treatment ofanaemia in animals and human beings, but also in any cases where aneasily absorbable source of iron is required.

The examples given below illustrate the process according to theinvention.

EXAMPLE 1 a. To 500 ml of an aqueous ferric chloride solution being 1.75molar as to iron, are added with vigorous stirring 500 ml of a 1.34molar aqueous sodium carbonate solution to form a ferric hydroxide sol.To the said sol are added 720 ml of a 25 percent aqueous solution ofdextran glycerol glucoside having an intrinsic viscosity of 0. l 8 at 25C. This corresponds to l .27 moles of anhydroglucose units per mole ofiron. Thereafter the solution is heated to 75 C and the temperature iskept at 75 C until the pl-l-value of the solution has decreased to 1. l.

The resultant iron dextran glucoside complex is precipitated from thesaid solution by adding 1,700 ml of methanol. The supernatant liquor isdiscarded and the complex is dissolved again in 600 ml of water at 50 C.The said process is repeated until the solution is mainly freed offerric ions. The solution resulting from the last solution in water isheated to C for 30 minutes to remove the methanol and is then heated for25 minutes in an autoclave at 1 10 C and at a pressure above theatmosphere amounting to 0.5 kg/cm". After cooling a 5 percent sodiumhydroxide solution is added until the pH-value of the reaction solutionis 11.3, whereafter heating for 20 minutes at 120 C at a pressure abovethe atmosphere amounting to 1 kg/cm is performed.

After cooling the pH-value of the solution is adjusted to 6.0 by adding4 N hydrochloric acid whereafter 100 percent methanol is added to obtaina methanol concentration of 36 per cent. Thereby is precipitated as afirst fraction 102 g of an iron dextran glycerol glucoside complex whichis isolated by decanting and dried. The precipitated product has an ironcontent of 27.2 percent and a dextran glucoside content of 48.5 percent.

To the decanted liquid 100 percent methanol is further added to obtain amethanol concentration of 60 percent. Thereby is precipitated as asecond fraction 120.6 g of ferric dextran glucoside complex containing 2percent of iron and 88 percent of dextran glycerol glucoside andcorresponding to 67 percent of the originally used dextran glycerolglucoside. The said fraction is used as starting material in anothercomplexing step.

26.2 g of the first precipitated fraction are dissolved in 80 ml ofwater. The solution is made isotonic with the blood fluids by addingNaCl. The final solution has a pH-value of 6.6, an iron content of mgper ml and an LD -value of 830 mg of Fe/kg weight of the body assayed byintravenous injection in mice.

b. To 500 ml of an aqueous ferric nitrate solution being 1.75 molar asto iron are added while stirring 500 ml of a 2.68 molar sodium hydroxidesolution to form a ferric hydroxide sol. To the said sol are added 200 gof the above mentioned iron dextran glycerol glucoside complexprecipitated as the second fraction, dissolved in 650 ml of water,whereafter the solution is heated to 50 C and this temperature is keptfor 45 minutes. After cooling the solution is dialysed with ionechangedwater for about 24 hours, whereafter it is heated in a sealed autoclavefor 15 minutes at 110 C. The solution is cooled again and its pH-valueadjusted to 10.8 by adding a 5 percent sodium hydroxide solutionwhereafter heating in a sealed autoclave for 30 minutes at 120 C takesplace. After cooling the pH-value is adjusted to 6.0 with 4 Nhydrochloric acid.

Then 100 percent methanol is added to obtain a methanol concentration ofabout 39 percent. Thereby is precipitated as a first fraction 95.2 g ofiron dextran glycerol glucoside complex which is isolated by decantingand dried. The dried product has an iron content of 31.5 percent and adextran glycerol glucoside content of 45 percent.

Further 100 percent methanol is added to the decanted liquid to obtain amethanol concentration of 60 percent. Thereby is precipitated as asecond fraction 110 g of iron dextran glucoside complex containing 2.5percent of Fe and 86 percent of dextran glucoside. The yield by weightis 55 percent of the iron dextran glycerol glucoside used in the secondcomplexing step. The said fraction is used as starting material inanother complexing step. 30.1 g of the first fraction are dissolved inml of water and the solution is made isotonic with the blood fluids byadding NaCl. The resultant solution contains mg of Fe per ml, has apl-l-value of 6.8, a viscosity at 25 C of 13 cps and an LD -value of1,150 mg of Fe/kg weight of the body assayed by intravenous injection inmice.

In the processes described above between 80 and 90 percent of theemployed dextran compound are utilized.

EXAMPLE 2 a. A ferric hydroxide sol is produced by adding 500 ml of a2.68 molar aqueous sodium hydroxide solution while stirring to 500 ml ofan aqueous ferric trichloro acetate solution being 1.75 molar as toiron. Then 1,000 ml of a 18 per cent aqueous solution of dextran havingan intrinsic viscosity of 0.35 at 25 C are added, and a complexingreaction as that mentioned in Example la is effected.

After the complexing step the solution is purified by ion exchange witha mixture of an acid and alkaline ion exchanger until the pH-value ofthe solution is 2.3, which indicates that the solution is mainly freedof ferric ions. Then the solution is heated in an autoclave for 30minutes at 120 C, cooled and treated with sodium hydroxide solution, asmentioned in Example la. After cooling 100 percent acetone is added toobtain an acetone concentration of 33 percent. The first fractionthereby precipitated is isolated, and 143 g of iron dextran complex areobtained, said complex containing 19.1 per cent of iron and 58 percentof dextran, which corresponds to 79.4 percent by weight of the dextranoriginally used.

Then 100 percent acetone is further added to obtain an acetoneconcentration of 55 percent. The supernatant liquor is decanted whereby63 g of iron dextran complex are obtained, said complex containing 0.1per cent of Fe and 90.2 percent of dextran and corresponding to 35percent by weight of the dextran originally employed.

b. 100 g of the last precipitated fraction are again subjected to thecomplexing reaction described in Example lb, the ferric hydroxide solbeing prepared from 230 ml of a 2.68 molar aqueous sodium hydroxidesolution and 250 ml of a ferric chloride solution being 1.75 molar as toiron. After the complexing step is completed, the resultant iron dextrancomplex is purified and precipitated by adding 1,000 m] of acetone. Theprecipitated complex is isolated by decanting and dissolved in 300 ml ofwater and the said process is repeated until the ferric ions beingpresent have been substantially removed.

After purification there is proceeded in the manner described in Examplelb, the fractionated precipitation being carried out by adding 100percent acetone to arrive at an acetone concentration of 34 percentfollowed by a further addition of acetone to attain an acetoneconcentration of 55 percent. The precipitated fractions are isolated anddried. 49.5 g of an iron dextran complex containing 30.3 percent of Feand 43 percent of dextran reckoned on the weight of the dried productare obtained as a first fraction and as a second fraction 52 g of aniron dextran complex containing 2.5 percent of Fe and 87 percent ofdextran reckoned on the weight of the dried product. The yield by weightof the two fractions is 49.5 percent and 52 percent, respectively, ofthe employed ferric dextran.

From the first fraction a solution is prepared as described in Examplelb, said solution containing 8 percent of Fe and having a viscosity of10 cps at 25 C and an LD -value of 890 mg of Fe/kg weight of the bodyassayed by intravenous injection in mice.

In the processes described above 70 to 75 percent of the employed amountof the dextran compound are utilized since only 100 g of dextran wereemployed. On industrial scale the utilization would have been better.

EXAMPLE 3 a. The first complexing step takes place in the same manner asdescribed in Example 1a, use, however, being made of 200 g of dextranglycerol glucoside having an intrinsic viscosity of 0.08 at 25 C.

After the complexing step the resultant iron dextran glycerol glucosidecomplex is precipitated by adding 2,000 ml of propanol-2 and theprecipitated product is isolated by decanting and dissolved in 600 ml ofwater. The said measure is repeated until excess of ferric ions has beensubstantially removed. After the last precipitation and filtering theproduct is dissolved in 1,200 ml of water and heated in a sealedautoclave for 10 minutes at 1 10 C whereafier there is proceeded asdescribed in Example 1a, with the exception, however, that thefractionated precipitation takes place at a propanol concentration of 35percent in precipitating the first fraction and a propanol concentrationof 55 percent in precipitating the second fraction.

Thereby are obtained as the second fraction g of iron dextran glucosidecomplex, containing 1.3 percent of Fe and 91 percent of dextranglucoside and corresponding to 45 percent of the employed dextranglucoside.

b. 90 g of the said fraction are used as starting material in a secondcomplexing step carried out as described in Example la, and after thecomplexing step the excess of ferric ions is removed by dialysis. Thenthe procedure described in Example lb is followed with the modificationthat the fractionated precipitation is carried out in a concentration of36 percent and then in a concentration of 60 percent. The precipitatedfractions are isolated and dried.

The first fraction amounts to 40.5 g of an iron dextran glucosidecomplex containing on the basis of the dry weight 30.5 percent of Fe and44 percent of dextran glucoside, and from this product there is preparedin the manner described in Example lb a solution containing mg of Fe perml and having a viscosity at 25 C of 14 cps and an LD -value of 890 mgof Fe/kg weight of the body assayed by intravenous injection in mice.

EXAMPLE 4 To 500 ml of a ferric hydroxide sol prepared as described inExample lb are added 385 g of a complex of ferric hydroxide withdextran, said complex containing 22 percent of Fe and 55 percent ofdextran, and having been prepared by the process described in Example 3a(1a). The amount of iron dextran used corresponds to 880 g per mole ofiron in the ferric hydroxide sol.

In other respects there is proceeded as described in Example lb, except,however, that the fractionated precipitation is carried out by addingacetone. In order to precipitate the first fraction 100 percent acetoneis used to obtain an acetone concentration of 34 percent.

After drying 244 g of an iron dextran complex are obtained containing32.7 per cent of Fe and 43 per cent of dextran. Therefrom is preparedexactly in the manner described in Example lb a solution containing 8percent of Fe and having a viscosity at 25 C of 9 cps and an LD -valueof 1,020 mg of Fe/kg of the body, assayed by intravenous injection inmice.

EXAMPLE 5 a. The first complexing step takes place in the same manner asdescribed in Example 3a.

b. 90 g of iron dextran glycerol glucoside complex containing 1.3% of Feobtained as the second fraction in Example 3a are used as startingmaterial in a second complexing step carried out as follows:

To 250 ml of an aqueous ferric chloride solution being 1.75 molar as toiron at 50 C are added with vigorous stirring 250 ml of a 1.34 molaraqueous sodium carbonate solution at 50 C to form a ferric hydroxidesol. To the said sol are added the 90 g iron dextran glycerol glucosidecomplex dissolved in 300 ml distilled water at 80 C. The temperature ofthe solution is kept at 65 C for 35 minutes.

The resulting iron dextran glucoside complex is precipitated from thesaid solution by adding 1,250 ml 87 percent isopropanol. The complex isfreed of ferric ions by repeated precipitations with 87 percentisopropanol and thereafter is dissolved in 300 ml distilled water. Thesolution is heated for 20 minutes in an autoclave at C. After cooling a5 percent sodium hydroxide solution is added until the pl-l-value of thesolution is 1 1.3, whereafter heating for 20 minutes at C in anautoclave takes place.

After cooling the pH-value of the solution is adjusted to 6.8 by adding4 N hydrochloric acid whereafter 87 percent isopropanol is added toobtain a final isopropanol concentration of 35 percent. The precipitatediron dextran glycerol glucoside is redissolved in 300 ml distilled waterand reprecipitated with 87 percent isopropanol to obtain a finalisopropanol concentration of 32 percent. Thereby is precipitated an irondextran glucoside complex which is isolated by centrifugation. Theanalysis of the dried product shows as follows:

Iron contents: 41.5%

Dextran glucoside: 21.3%

Moles iron per anhydro- 5.66 Fe/anhydroglucose unit: glucose H O: 2.1%

Intrinsic viscosity: 0.07

Contents Dextran/Contents Fe: 0.51

28.6 g of this iron dextran glucoside complex are dissolved in 90 mldistilled water. The solution is made isotonic with the blood fluids byadding NaCl. The final solution has a pH-value of 7.0, an iron contentof 120 mg per ml, a viscosity at 25 C of 15 cps, and an LD -value of1,050 mg of Fe/kg weight of the body assayed by intravenous injection inmice.

EXAMPLE 6 a. The first complexing step takes place in the same manner asdescribed in Example 1a, but the amountsused are upscaled 100 times.

b. 7.2 kg of ferric dextran glucoside complex containing 2 per cent ofiron and 88 percent of dextran glucoside obtained as the second fractionin the above mentioned Example are used as starting material in thesecond complexing step carried out as follows:

To 20 liters of an aqueous solution containing 9.4 kg of ferric chloridehexahydrate at 45 C are added with vigorous stirring 20 liters of anaqueous solution containing 3.6 kg of anhydrous sodium carbonate at 45 Cto form a ferric hydroxide sol stabilized by excess of chloride ions. Tothe said sol 7.2 kg of ferric dextran glucoside complex dissolved in 30liters of deionized water at 85 C are added. The temperature of thesolution is kept at 60 C for 30 minutes.

The resulting iron dextran glucoside complex is freed offerric ions byrepeated precipitations with 99 percent isopropanol and thereafterdissolved in 30 liters of deionized water. The solution afterevaporation of the isopropanol residue is heated in an autoclave for 20minutes at 100 C. plus 15 minutes at 1 10 C.

After cooling to 50 C a 10 percent sodium hydroxide solution is addeduntil the pI-l-value of the solution is 11.0, whereafter heating forminutes at 120 C takes place.

After cooling to 60 C the pI-I-value of the solution is adjusted to pH6.5 with hydrochloric acid, whereafter 99 percent isopropanol is addedto obtain a final isopropanol concentration of 36 percent.

The precipitated iron dextran glucoside complex is redissolved in 30liters of distilled water and precipitated fractionally with 87 percentisopropanol to obtain a final isopropanol concentration of 33 percent.The precipitated iron dextran glucoside is isolated by centrifugation ina Super Centrifuge, redissolved in 25 liters distilled water andspraydried. A total amount of 4.0 kg dry powder is obtained the analysisof which shows as follows:

Iron contents: Dextran glucoside:

37.8 percent 27.3 percent With the dry powder an isotonic solution ismade with an iron content of 100 mg per ml, a viscosity at 25 C of 8 cpsand an LD -value of 1,200 mg of Fe/kg body weight when assayed byintravenous injection in mice.

The eluate of the centrifuge containing the iron dextran glucoside withlow iron content is precipitated with 99 per cent isopropanol to a finalconcentration of 60 percent isopropanol. After drying 5.8 kg of ferricdextran glucoside complex containing 2.9 percent of iron and percent ofdextran glucoside are obtained. This is a recovery of percent based uponthe initial dextran glucoside.

In the process of the present invention the loss of dextran compound isconsiderably lower than in the processes known in the art. In Examplesla, lb and 2a the dextran glucoside loss is only between 10 and 20percent. In Example 2b the loss is a little higher due to the fact thatthe process is carried out on a very small scale, only using g ofdextran glucoside in the complexing procedure. When carrying out theprocess of the invention on an industrial scale the loss of dextrancompound is less than 10 percent, vide example 6.

In Example 3 of the U.S. Reissue Pat. specification, No. 24,642 thereare used 400 g of dextran as starting material whereas the final product600 ml liquid contains 5 percent of Fe. Nothing is said about thedextran content in the final product, but in the specification it issaid in col. 3, lines 39 to 40 that a preparation containing 5 percentof Fe will contain 30 to 50 percent of dextran. If the preparationcontains 30 percent of dextran the 600 ml liquid will contain g ofdextran corresponding to a dextran loss of 55 percent. If thepreparation contains 50 percent of dextran the 600 ml liquid willcontain 300 g of dextran corresponding to a dextran loss of 25 percent.

It must be considered that preparations containing 5 percent of Fe and30 percent of dextran in the solution will contain about 12.5 percent ofFe and 75 percent of dextran in dry state while preparations containing5 percent of Fe and 50 percent of dextran in the solution will containabout 9 percent of Fe and 80 percent of dextran in dry state. Of theselarge amounts of dextran only a small fraction is complexed to iron. Therest of the dextran (as much as 80 percent) is unbound and could havebeen utilized for complexing more iron. In the Example of the Swiss Pat.specification, No. 370,194 the starting materials are 50 g ofpolyisomaltose and 80 ml of an aqueous solution containing 30 percent byweight of FeCl ,6I-I O, which is equal to 5 g of Fe. The final productis an iron/polyisomaltose complex containing 33.4 percent of Fe. If

it is supposed that all the 5 g of Fe were complexed to thepolyisomaltose, which is the most favorable condition for the reaction,and because of the fact that the final product according to theindications in the specification, vide page 2, lines 2 to 6, shows theratio 2 moles of Fe to each anhydroglucose unit of the polyisomaltose,the following amount of polyisomaltose was complexed to iron:

5 X 162/2 X 55.8 7.3 g of polyisomaltose This means that 50 g minus 7.3g 42.7 g of polyisomaltose have not been complexed. The loss ofpolyisomaltose is thus more than 80 percent.

In the Example of U.S. Pat. specification, No. 2,885,393 the loss ofdextran is about 60 percent. In the Example there are used 90 g ofdextran and the final product is 82 g of the iron dextran complexcontaining 24 percent of Fe, which means OH-groups about 20 percent. Thecontent of dextran may be calculated as follows: 100 minus 24 minus 2056 percent. The 82 g of iron dextran complex thus contain 5 6 X 82/10046 g of dextran and this corresponds to a dextran loss of about 60percent.

The above comparison shows that in our process it is possible to obtainhigher Fe content in the iron dextran complex and that the loss ofdextran compound is considerably lower in our process than in the knownprocesses referred to above.

The purpose of making iron dextran complex is to bring iron into anon-toxic form and to make it available to the organism by injection.The purpose of the production of iron dextran complexes is not to supplythe body with dextran. This means that it constitutes an improvement inthe art to make available to the public an iron dextran complexpreparation with a high iron content and a low dextran content.

What we claim is:

1. In a process of producing complexes of ferric hydroxide and a dextrancompound having an intrinsic viscosity within the range of 0.025 and0.75 at 25C and selected from the group consisting of dextran anddextran glucosides, the improvement wherein the complexing reaction iscarried out in two steps and wherein a fraction of the complex from thefirst complexing step is used in the second complexing step 2. Themethod of increasing the iron content of iron dextran complexes whichcomprises reacting an iron dextran complex in an aqueous medium underacid conditions with a formed in situ ferric hydroxide sol andthereafter precipitating less than about one half the iron dextrancomplex from the reaction mass and separating and recovering thefraction precipitated.

3. The method according to claim 2 wherein a second fraction isprecipitated separated and recycled.

4. The process for producing iron dextran complexes which comprises afirst complexing step in which ferric hydroxide is reacted under aqueousacid conditions with a dextran compound having an intrinsic viscosity of0.025 to 0.75 at 25 C and selected from the group dextran and dextranglucosides, precipitating the iron dextran complex into at least twofractions having different iron contents, subjecting one of thefractions of the iron dextran complex formed in the first complexingstep to a second complexing step and wherein said iron dextran complexis reacted under aqueous acid conditions with ferric hydroxide,precipitating the iron dextran contained in the reaction mass into atleast two fractions having different iron contents, separating andrecovering the fraction having the higher iron content and recycling thefraction having the lower iron content.

5. A process according to claim 4 wherein the fraction of the irondextran complex formed in the first complexing step that is subjected tothe second complexing step is the fraction having the lower ironcontent.

6. A process according to claim 4 wherein both of the complexing stepsare carried out at a pH of l. l to 2.3.

7. A process according to claim 4 wherein unreacted ferric ions areremoved from the complex produced in the first complexing step before itis subjected to the second complexing step.

8. A process according to claim 4 wherein an aqueous alkaline solutionof the iron dextran complex from the first or second complexing steps isautoclaved, cooled and acidified before fractionally precipitating theiron dextran complex.

9. A process according to claim 4 wherein an aqueous alkaline solutionof the iron dextran complex from the first and second complexing stepsis autoclaved, cooled and acidified before fractionally precipitatingthe iron dextran complex.

2. The method of increasing the iron content of iron dextran complexeswhich comprises reacting an iron dextran complex in an aqueous mediumunder acid conditions with a formed in situ ferric hydroxide sol andthereafter precipitating less than about one half the iron dextrancomplex from the reaction mass and separating and recovering thefraction precipitated.
 3. The method according to claim 2 wherein asecond fraction is precipitated , separated and recycled.
 4. The processfor producing iron dextran complexes which comprises a first complexingstep in which ferric hydroxide is reacted under aqueous acid conditionswith a dextran compound having an intrinsic viscosity of 0.025 to 0.75at 25* C and selected from the group dextran and dextran glucosides,precipitating the iron dextran complex into at least two fractionshaving different iron contents, subjecting one of the fractions of theiron dextran complex formed in the first complexing step to a secondcomplexing step and wherein said iron dextran complex is reacted underaqueous acid conditions with ferric hydroxide, precipitating the irondextran contained in the reaction mass into at least two fractionshaving different iron contents, separating and recovering the fractionhaving the higher iron content and recycling the fraction having thelower iron content.
 5. A process according to claim 4 wherein thefraction of the iron dextran complex formed in the first complexing stepthat is subjected to the second complexing step is the fraction havingthe lower iron content.
 6. A process according to claim 4 wherein bothof the complexing steps are carried out at a pH of 1.1 to 2.3.
 7. Aprocess according to claim 4 wherein unreacted ferric ions are removedfrom the complex produced in the first complexing step before it issubjected to the second complexing step.
 8. A process according to claim4 wherein an aqueous alkaline solution of the iron dextran complex fromthe first or second complexing steps is autoclaved, cooled and acidifiedBefore fractionally precipitating the iron dextran complex.
 9. A processaccording to claim 4 wherein an aqueous alkaline solution of the irondextran complex from the first and second complexing steps isautoclaved, cooled and acidified before fractionally precipitating theiron dextran complex.